Telomerase Reverse Transcriptase Gene Transfection of Human Amniotic Electrical Mesenchymal Stem Cell Transplantation for Treatment of Diabetes Mellitus

ęඈඋ1࣏ۘऺ ৰ 19 Ż ৰ 36 ƛ 20150903 ϦČ Chinese Journal of Tissue Engineering Research September 3, 2015 Vol.19, No.36 www.CRTER.org

঳ୖ›ǑḰơ›ȖüϹḰɏ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉ͏ћசɳ҉

Äņᇽ(ǧSďS˂ˡŋʒ½ˍãKǧSď 300350)

তͥ fK˿K1983 ˝ÛK ĚȈ࿁3Ⴎu̩:࿁ṇƆ਍Ľͥ «ֲQ¾ඊྡྷ͹ϣۘऺ:ǻₑᡅՈඊྡྷࢿΉ ȩ ၠⒸ̱ᯬ¾ඊྡྷͣƅ̶๎ ǡļ7P4ত඗Șᜬ Ởẋ঳ୖ›ǑḰơ›ȖüϹḰɏ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉǳƅά͏ћசɳ҉4 ̂ZàǁďNK˩ȂK 2010 ˝ 7 ÅǧSãî͸ ͟⏲᪑ ϤKœþKå ۔ඊྡྷࢿΉ¦๎ၠⒸ̱ᯬ¾ඊྡྷசɳ҉ाွᜄசྴˏ௤঳ୖ›ǑḰơ›Ȗüᜄச֕ϟ̓¾ 'L᪑ ȁ·ȔŀYȩ fKǧS ¾ඊྡྷඈඋ1࣏ᜄசசɳ҉ ď S ˂ ˡ ŋ ʒ ½ ˍ ෭Щ᪱ ãKǧSď 300350

঳ୖ›ǑḰơ›human telomerase reverse transcriptase hTERT Ě ିǻ:R394.2 B:ۅǲʃ᪊ ʼᡅ তේǻ:2095-4344 ྐë๎ၠ«ྒ̫ϦϣȒՈëŏĭ ๎ၠⒸ̱ᯬ¾ඊྡྷͣƅǚǌ5ƫ3ʺȪ࿁Ɔ3\Ēˊƚuᩲ3̹ѯƣ (2015)36-05827-06 ɳĺ਍ĄÃ4 ɘ4ěY2015-07-25 http://WWW.crter.org ՈỞẋ঳ୖ›ǑḰơ›(human telomerase reverse transcriptase hTERT)ȖüϹḰɏ¦๎ၠⒸ̱ᯬ¾ඊֲ Ոћά4۔சɳ҉µƧ Ȇᩬ͢ȭசɳ҉̓۔ࢿΉ4̓ྡྷ ƶǚ╓۔̓ ࡿȕͧ¦๎ၠⒸ̱ᯬ¾ඊྡྷ ඓ hTERT ȖüϹḰɏ๎ၠⒸ̱ᯬ¾ඊྡྷ º 50 Ǯ SD 5ͩ

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ა[☝࿍ͼȸࢿΉ¦๎ၠⒸ۔/Ởẋ̓ ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈʐ hTERT¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈ๎¦ ᯬ¾ඊྡྷʐඓ hTERT ȖüϹḰɏ๎ၠⒸ̱ᯬ¾ඊྡྷ4ࢿΉȒȈඈẟᜐ¬ɍᜄசˈ¿֕ϟ zࢿΉȒɣɬ̼̱

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Ήඈᜄசˈ¿º[┑(P < 0.05) ɘ͢« hTERT¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈՈाွᜄசˈ¿ȉẕzȭ+ඈ ˈ¿(P > 0.05) ໐சɳ҉ඈाွᜄசමťńṇʌˈ¿ࢿΉȒ 6 ɬ ^சɳ҉ඈɨṇ ¦๎ၠⒸ̱ᯬ¾ඊྡྷ

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சɳ҉4۔ᜄசʐλḿྴˏǃĐ ǳÑƅά͏ћ̓۔ඊྡྷࢿΉ࿁º┑ĺசɳ҉̓

:(Äņᇽ. ঳ୖ›ǑḰơ›ȖüϹḰɏ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉ͏ћசɳ҉[J].ęඈඋ1࣏ۘऺ 2015 19(36 5827-5832. doi:10.3969/j.issn.2095-4344.2015.36.016

Telomerase reverse transcriptase gene transfection of human amniotic electrical mesenchymal stem cell transplantation for treatment of diabetes mellitus

Fu Jian-ru (Department of Internal Medicine, Xianshuigu Hospital of Jinnan District, Tianjin 300350, China)

Abstract BACKGROUND: The amniotic membrane is the rejected material after birth. Amniotic mesenchymal stem cells are characterized as easy harvesting, strong proliferation ability, no ethical controversy, and low immunogenicity. OBJECTIVE: To electrotransfer human telomerase reverse transcriptase (hTERT) gene into amniotic mesenchymal stem cells transplanted into diabetic rats and to explore its effect on diabetic rats. Fu Jian-ru, Master, Physician, METHODS: Human amniotic mesenchymal stem cells were isolated, cultured and electrotransferred by hTERT Department of Internal gene. Ten of 50 Sprague-Dawley rats were randomized selected as controls, and the remaining rats were used to Medicine, Xianshuigu Hospital establish diabetic models through injection of 45 mg/kg streptozotocin. Thirty-six model rats were randomized into of Jinnan District, Tianjin model group, cell transplantation group and hTERT-transfected cell transplantation group, with 12 rats in each 300350, China group. In the latter two groups, human amniotic mesenchymal stem cells and hTERT-transfected amniotic Corresponding author: Fu mesenchymal stem cells were injected via sublingual veins, respectively. After transplantation, blood glucose Jian-ru, Department of Internal levels were monitored dynamically, and plasma insulin concentration was detected every week. Pancreas tissues Medicine, Xianshuigu Hospital were taken and cut into sections for histological observation using hematoxylin-eosin staining. of Jinnan District, Tianjin RESULTS AND CONCLUSION: At 4 weeks after transplantation, the blood glucose levels were significantly lower in 300350, China the two cell transplantation groups than the model group (P < 0.05), and especially in the hTERT-transfected cell transplantation group, the blood glucose level was close to the normal value (P > 0.05). However, the model group still Accepted: 2015-07-25

ISSN 2095-4344 CN 21-1581/R CODEN: ZLKHAH 5827

Äņᇽ. ঳ୖ›ǑḰơ›ȖüϹḰɏ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉ͏ћசɳ҉ www.CRTER.org had a higher blood glucose level. At 6 weeks after transplantation, compared with the model group, the plasma insulin concentration was significantly increased in the two cell transplantation group (P < 0.05), and the severity of pancreatic injury was also eased in these two groups (P < 0.05), especially in the hTERT-transfected cell transplantation group (P < 0.05). These findings indicate that hTERT-transfected amniotic mesenchymal stem cell transplantation can dramatically decrease blood glucose level and relieve pancreatic injury in diabetic rats, which is an effective method for treatment of mellitus diabetes in rats.

Subject headings: Stem Cells; Tissue Engineering; Blood Glucose; Diabetes Mellitus

Fu JR. Telomerase reverse transcriptase gene transfection of human amniotic electrical mesenchymal stem cell transplantation for treatment of diabetes mellitus. Zhongguo Zuzhi Gongcheng Yanjiu. 2015;19(36):5827-5832.

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Äņᇽ. ঳ୖ›ǑḰơ›ȖüϹḰɏ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉ͏ћசɳ҉ www.CRTER.org zა[☝࿍ͼȸ45 mg/kgFₓՈ⏂࿶◍௤(ϬQŢzpH ⒸϔǒɨṇϬʳƨt ̼ɀ ̼ɀˈβα=0.054 4.5Ոɜŀ‼֔෗Ξ^) 1ɬͱẢන2Ƶ3Ⓒ╘ႷɅ2 d Ƨ ƵͼȸȒ zɣɬǚɲ☝࿍ᜄச1Ƶ ᜄச֕ϟẢන3Ƶ̓ 2඗ȘResults &50Ǯ ╓ƶǚ10Ǯň۔z16.7 mmol/L ໐/Ϧɴ̶3̶Ģ3̶ɳ3ĿᯬₓλɅ 2.1 ôκŦʦļƘǫʭ ൷͑̓ ȭ+ඈ 36Ǯǆ^Ụµ ȏʐͫɏ4ǮX┨ żඌᩥ͑ ۔Ụµt£4Ụµẋ࣏ 3Ǯ¬ĭȏ 1ǮƌϏՈ̓& ᜄசϧò]ࣷǎ tÑX┨ żඌỤµt£¬ĭϔ&36Ǯ4 ඗Ș ȌՈϔₓ46Ǯ4 ň&ࢿΉǛĿ4Ⱥசɳ҉ 2.2 ̱ωĝσʗϏƉȼˍFĝ́Ƨ ᰻!ȕͧՈ๎ၠ ۔Ⱥசɳ҉Ụµt£Ո36Ǯ̓ &hTERT_¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈ3¦๎ Ⓒ̱ᯬ¾ඊྡྷɌ᯸˝ϣ⑃ Ʈɍ̶ʳ Ɍ̩Ʈ3̶ṽƮ3 ƶ╓۔̓ ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈʐசɳ҉ඈ ɣඈȈ12Ǯ hTERT_ Ʈ3ൾ⏨Ʈz̶ᢖƮ਍(Ě1A)4ČႷৰ2ÏȒ๎ၠⒸ̱ᯬ ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈ3¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈ ¾ඊྡྷ᯸˝ợò¤ȷ 8 hό:ǀ͔᯸˝ ඊྡྷɌɴţĄP ა[☝࿍ͼȸࢿΉ๎ၠⒸ̱ᯬ¾ඊྡྷ˸^/ ႸՈt൨මඊྡྷʳ ඊྡྷ̶Ɍřɶ΢ȸƮź3⑃̩ƮzG۔Ởẋ̓/ hTERT๎ၠⒸ̱ᯬ¾ඊྡྷ˸^ ¦๎ၠⒸ̱ᯬ¾ඊྡྷ˸ ॡźϣ⑃ ǾาºǺ␠[ᢆȓ4 ๎ၠⒸ̱ᯬ¾ඊྡྷ¿ţ ȯƅ਍ₓ ȀՈⒸՈඊྡྷϧò&2×106 L 14 ָǐ൪&15 µm ྡྷϚͱȯƅ^ 1.4.4 RT-PCR̼ϟ ỞẋTRIzolͩȴǚʇRNA ₋Ϭ௯ ՈʌɈȖ̩ȌĿ3ඃୖĿʐႸǺĿ ྡྷϚɬĐƅႮा͵ òᩥϟǎRNAȯₓ ÑǑḰơȌt᪙F֖᪸R ʐྡྷĢɃ͵ ඊྡྷzඊྡྷॅ᰻7Ⓒǳᢅ4ඊྡྷẢȉ඗Ȁ̵̵ ̲ ẟᜐǑḰơ4PCRǑàǝâ94 ?ǜɳ5 min 95 ?ǜ ᢅĚ1B4ȕͧՈ๎ၠⒸ̱ᯬ¾ඊྡྷඓϕśඊྡྷÖℸǎȒǳ ɳ30 s 54 ?Ễ̗36 s 72 ?łĤ30 s 36ɬƛǶɳȒ ᜬẂּàՈʃᩴĭ ܲᩨȕͧඊྡྷ&๎ၠⒸ̱ᯬ¾ඊྡྷ4 72 ?łĤ6 min4z4 ?[ǉƌĭ4PCRɣPǑàₑ̩ 2.3 hTERTΆ9ǇA̢ÝσʗϏƉȼˍFƏĝ2ê ẟᜐ3Ƶ4PCRĭǚ5 mLẟᜐϹ· Ñβ-actinň&ͱǆ RT-PCR̼ϟº߾ ϹḰɏͩ·ȰhTERTȖüḰɏ๎ၠⒸ̱ +4Ϲ·ȒาGDSS000ωྺႮ¬tʻÖZı+ǉƌĚʻ ᯬ¾ඊྡྷ48 hȒ ඗Șº߾Ϧ2ǝ±ìǝr ᜬRNAᯬₓ

àϬImage-Pro Plus 8.0ḳâ ȌhTERTՈǝr^β-actin Ι -Ϭ௯̲ඃ ̵̵òÖ̼ϟRNAńA260 nm A280 nm̠

Ȍ4 Ոȼ̵òȨ ඗Șº߾A260 nm/280 nm=1.82.0 ᜬRNA൳ ǝrՈּȭ̜òɨȨ ẟᜐmRNAŎǎₓ òṇΙ PCRĭ̼ϟℸǎ඗ȘᜬhTERT_๎ၠⒸ̱ šĭÛ ᯬ¾ඊྡྷḰɏඈń750 bpǳᢅĽŎɳǝr ໐¦๎ၠⒸ̱ Ȗü šĭÛ ċɉ⑃ò ᯬ¾ඊྡྷࢿΉඈ3சɳ҉ඈ̼ϟƦᢅĽŎɳǝrᜬ\ hTERT Zä5’-CTGAAGTGTCACAGCCTGTTT-3' 1 1 1 bp hTERT mRNAՈᜬẂ ᢅĚ24 [ä5’-CACACATGCGTGAAACCTGTA-3' 1 1 1 bp 2.4 σʗϏƉȼˍF ű̢ɪȆîĲͻǂΛͻ ē β-actin Zä5’-TATCGGACGCCTGGTTAC-3' 151 bp ŵŋͬ)̼ ࢿΉȒ4ɬ ȭ+ඈᜄசǉť൪4.0 mmol/L [ä5’-CTCAGCCTTGACTGTGCC-3' 1 51 bp சɳ҉ඈᜄச2226 mmol/LǳᢅºŋʌhTERT¦๎ h TERT঳ୖ›ǑḰơ›4 ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈᜄசǉť൪7.01 mmol/L3¦๎ၠⒸ / ᜄசǎᜄ±ྴ ̱ᯬ¾ඊྡྷࢿΉඈᜄசǉť൪12.03 mmol/Lţƅ┑ĺ۔ၠⒸ̱ᯬ¾ඊྡྷࢿΉȒȈඈ̓๎ 1.4.5 ɣ hTERT¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈṇ¦๎ၠⒸ̱ᯬ¾ඊ۔௤ˈ¿̼ϟ ๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉȒ ȭȈඈ̓ˏ ɬǎrzɲ☝࿍₋ᜄ -ϬᜄசÖർċͩϟǎाွᜄச ྡྷࢿΉඈՈᜄச┑ĺŰº(P < 0.05) ᢅᜬ14 ɣɬ2Ƶ4Ảන֕ϟᜄச6ɬ4z๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉȒ ࢿΉȒ6ɬ ^சɳ҉ඈɨṇ ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿ Ȓඓွ'¬ ΉඈʐhTERT¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈՈᜄϚྴˏ௤⁍ٯ6ɬ ₋Ϭ7%ˈȌ˃ (5 mL/kg)ွဘͼȸ ࿍ǚᜄ ࡿȏ Ñ20 ?ǉНᜄ± ƉǒELISA᪙F֖Ẕ ˈ¿ʺ¤ ྴှ҉ǃ࣏òλḿ ^¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿ ᜄϚྴˏ௤ˈ¿4 Ήඈּɨṇ hTERT¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈŰº۔ň᪸R ̼ϟȈඈ̓ 1.4.6 ๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉȒ҉ˊ̼ϟ ๎ၠⒸ̱ᯬ (P < 0.05) ᢅᜬ24 ¾ඊྡྷࢿΉȒৰ6ɬ ඓွ'¬࿍ǚᜄȒ ẉợ-Ϭ)ʇͩ 2.5 σʗϏƉȼˍF ű̢îĲēȆŮDŨ)̼ ඈඋƅ]Ȑ࣏ˏྴ۔ှඈඋ Ñ40 g/L̶༞϶ Ėǎ ๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉȒ6ɬ Ȉඈ̓ྴ۔ȏ¬ĭ ǚȈඈ̠̓ ˏྴ ċ ₋ϬᆓƤஂö൦ᢈɏᄶ ᢆȓ̓ òՈǃǧ ྴˏඊྡྷūȏ ྴˏṽො̺ɳඊྡྷ$R ᗥẢනڷ ẋ̸ ๎¦Ո҉ˊǜĚ4 ₑ۸ū ྴˏඊྡྷᢅɅ 3ඈּɨṇ໐ᣄ hTERTˏྴ۔ 1.5 ƶϙʬ+ŷ/ ᢆȓȈඈᜄச¬ɍˈ¿ǜĚ zࢿΉ ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈՈྴˏඊྡྷǑàǳᢅºλḿ / ċᆓƤஂ_ ඗ȀǀϘ ྴˏඊྡྷṇ̶ ࢿΉȒ6ɬ ^சɳ҉ඈɨṇ ᜄϚྴˏ௤Ոϧò ྴှ۔Ȓɣɬ̼ϟȈඈ̓ ö൦ɏᄶᢆȓ҉ˊǜĚ4 ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈʐhTERT¦๎ၠⒸ̱ᯬ¾ඊ 1.6 Ȍα͸ǫʭ ǒɀϔǒţ₋ϬSPSS 15.0ḳâிඣ ྡྷࢿΉඈྴှ҉ǃ࣏òλḿ ɘ͢«hTERT_¦๎ၠⒸ̱ _ ẟᜐ Ȍ ϔȨ৪ȌǷɍ O ᩥₓᰈÑx±sᜬ߾ ඈ ᯬ¾ඊྡྷࢿΉඈŰº(P < 0.05) ᢅĚ34

ISSN 2095-4344 CN 21-1581/R CODEN: ZLKHAH 5829

Äņᇽ. ঳ୖ›ǑḰơ›ȖüϹḰɏ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉ͏ћசɳ҉ www.CRTER.org

A B Ě 1 ȕͧՈ๎ၠⒸ̱ᯬ¾ඊྡྷՈƮɍ(Ǿา

ºǺ␠) Figure 1 Morphology of amniotic mesenchymal stem cells cultured under inverted microscope ĚͼĚ A &᰻!ȴǚՈ๎ၠⒸ̱ᯬ¾ඊྡྷ (×40)Ɍ⑃̩ƮB &ČႷৰ 2 ÏȒՈ๎ၠⒸ̱ ᯬ¾ඊྡྷ ɌɴţĄPႸՈt൨මඊྡྷʳ඗Ȁ (×100)4

1 2 3 4

Ě 2 ঳ୖ›ǑḰơ›(hTERT)ȖüḰɏȒń๎ၠⒸ̱ᯬ¾ඊྡྷՈ ᜬẂ bp Figure 2 hTERT expression in amniotic mesenchymal stem cells

2 000 after transfection Ěͼ14 & Marker ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈ3சɳ҉ඈ3 1 000 hTERT2๎ၠⒸ̱ᯬ¾ඊྡྷḰɏඈ4hTERT๎ၠⒸ̱ᯬ¾ඊྡྷḰɏ 750 ඈń 750 bp ǳᢅĽŎɳ mRNA ǝr4 500

250 100

A B C D

(ඈඋ҉ˊǜĚ(ᆓƤஂö൦ɏᄶ ×100ˏྴ۔̓ Ě 3 F igure 3 Histological changes of the pancreas islet of rats (hematoxylin-eosin staining, ×100) Ě ͼĚ A &ȭ+ඈǷྴˏඊྡྷB &சɳ҉ඈ ྴˏඊྡྷᢅɅ ̺ɳඊྡྷ$R ྴˏඊྡྷₑ۸ūC &¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈ ̺ɳ$RඊྡྷǑàλḿD & hTERT¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈ ྴˏඊྡྷǑàºλḿ ඗ȀǀϘ4ੱ͐¤ūÏᜬྴˏඊྡྷ4hTERT঳

ୖ›ǑḰơ›4

௤ˈ¿ǜĚˏྴ۔ᜄசǜĚ ᜬ 2 ๎ၠⒸ̱ᯬ¾ඊྡྷა[☝࿍ࢿΉȒȈඈ̓۔ᜬ 1 ๎ၠⒸ̱ᯬ¾ඊྡྷა[☝࿍ࢿΉȒȈrⒸɉȈඈ̓ _ _ (x±s mmol/L) (x±s µg/L) T able 1 Variation of blood glucose levels in rats at different time after Table 2 Variation of fasting insulin levels in rats at different time a mniotic mesenchymal stem cell transplantation via sublingual veins after amniotic mesenchymal stem cell transplantation via sublingual veins ඈ / n 5 d 14 d

ȭ +ඈ 10 4.01±0.05a 4.02±0.01a ඈ/ ࢿΉQ ࢿΉȒ6ɬ ab ab ச ɳ҉ඈ 12 22.35±0.06 22.42±0.04 ȭ+ඈ 14.61±5.32ab 15.41±3.02ab ab ab ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈ 12 20.03±0.32 19.68±0.01 சɳ҉ඈ 0.33±0.05 0.30±0.06 b b hTERT¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈ 12 17.01±0.12 14.27±0.01 ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈ 0.28±0.03ab 9.02±0.11ab a a ඈ/ n 21 d 28 d hTERT¦๎ၠⒸ̱ᯬ 0.29±0.10 12.38±0.32 ¾ඊྡྷࢿΉඈ ȭ+ඈ 10 4.02±0.04a 4.02±0.01a a ச ɳ҉ඈ 12 23.67±0.07ab 26.21±0.08ab ᜬͼ^சɳ҉ඈɨṇ P < 0.05^ hTERT¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈɨ ab ab b ¦ ๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈ 12 14.32±0.03 12.03±1.08 ṇ P < 0.054hTERT঳ୖ›ǑḰơ›4 hTERT¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈ 12 9.08±0.02b 7.02 ±0.07b

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3ᩬ᩾Discussion ᯬ¾ඊྡྷࢿΉȒ࿁̻ƯūǃĐՈྴˏඈඋඊྡྷʺȪ ẟ໐ ǂ˥DĝƍƟǒƽĝ˫Ǒ˼̵řǮ ֲQȭzசɳ Ưūᜄச[┑4 3.1 Ո͏ћ5ͩ ẜɎƦƅǳÑẂ4Ǉá͏͔Ո5ͩ4ɴ Ñǌȭz๎ၠⒸ̱ᯬ¾ඊྡྷՈۘऺţᜬ๎ၠⒸ̱ᯬ҉ ⓺ɉ₋ǚՈ┑சሳĭǎɬ௤ћͩKƦ࿁Ǉá͏͔[14-16] ¾ඊྡྷ«·zྞྒ¾ඊྡྷ^tĿ¾ඊྡྷⒸՈPିⒸ਍൫ ÖĿTØȺྴˏࢿΉň&͏͔சɳ҉Ոˊ̿5ͩ Ĳ«ƅ Ո¾ඊྡྷ üǸͣƅĺ̹ѯƣɳ3ʌʺȪ࿁3Ɔ ĚȈ ͢ǡļʐ̹ѯǑàՈ└:໐ƽˑrྴˏࢿΉՈàϬ4 ࿁3ǡļ5ƫ਍ưĽՈĄᱎɳ[32-37]4ĲŸ෾Ʌ๎ၠⒸ̱ᯬ ĚǺɳʟՈּۘ͟ऺ Kƽ ẕÀǡ ╓؄¾ඊྡྷࢿΉǒɀǎ ÖۘऺՈǕʉ ¾ඊྡྷȖü᫇ȋƶ:ǎǎȕ Ěάɋ ໐ǒɀ඗Șȴ சɳ҉ՈǇá͏͔rǡrXƗ4ϬǡۘऺՈ¾ඊྡྷΞྴ ˑr๎ၠⒸ̱ᯬ¾ඊྡྷՈʺȪʐ& ှ¾ඊྡྷ3᪵Ȱɳ̶࿁¾ඊྡྷÑǎ]ȐǡļՈⒸ̱ᯬ¾ඊ ߾ ඓhTERTȖü᫇ȋȒǳÑƯẟ๎ၠⒸ̱ᯬ¾ඊྡྷՈʺ ྡྷ਍[17-18] ͢¦๎ၠⒸ̱ᯬ¾ඊྡྷͣƅĺ̹ѯƣɳ3ʌ Ȫάɋ ࿁ƯūඊྡྷˌϣĚ4 ĚȈ࿁3ǡļ5ƫ਍ưĽĄÃ[19]4ƅۘऺ වZ¤Ẵ ỞẋhTERTϹḰɏ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿ Ȫ࿁3Ɔʺ Ǖɴ¦๎ၠⒸ̱ᯬ¾ඊྡྷńĿ̲᪵Ȱǝâ[ǳ Ětྴˏ Ήǳƅά͏ћசɳ҉4ʇ໐ᣄ7 ๎ၠⒸ̱ᯬ¾ඊྡྷͣƅ ĚȈ࿁3Ⴎu̩:࿁ṇƆ਍Ľͥ «ֲQ¾ඊྡྷ͹ ௤ϣඊྡྷ ÂǳƯẟᜄசʮ̩ႷǷˈ¿[20-22]4ẕÀ ̶ ǡ ̓ₓۘऺ>ᜬr๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉȭசɳ҉ͣ ϣۘऺ:ǻₑᡅՈඊྡྷࢿΉǡļ7P4Ĳüǒɀۘऺᰈ Ě£࿁ÑǎĿͱ ƅṇΙћά Ĳ«͢ňϬƶ:ƅǑẟPǹۘऺ[23-25]4 ƅ└ ͟z๎ၠⒸ̱ᯬ¾ඊྡྷՈ̶ȕ 3.2 hTERTĝjļΛǇAhTERT̢ˍFĝją hTERT ƌϏ3ẅࢿ3 Ěʐ^ǳ'ǌՈ඗ȀϘȌ3£࿁ϘȌẜ ঳ୖ›ՈʘĚřĹ ۘऺǕɴš͑hTERTǳÑūඊྡྷ ◄ᡅẟPǹ͑ۘऺȆᩬ4» ǉťǷᜬƧǎ ĚĽͥ ࿁ūš͑hTERTՈඊྡྷ঳ୖ⑃ òǣÑł⑃ ƯūඊྡྷቻǣˌϣĚ4ḰɏrhTERTՈ๎ၠ ň້ᯥǲôκÉαôèͯʑǧƨĔΚfɻΛÔ̖ Ⓒ̱ᯬ¾ඊྡྷǉťr๎ၠⒸ̱ᯬ¾ඊྡྷՈĽͥ ͣƅࣷǎ ¸ɡȩ f ঳ୖ›ϏɳᜬẂ ÂᜬɴϦࣷǎťනʺȪՈĄÃ4hTERT -֎ΞॅƖuȔŀʃȅRæɻģʕ˵ʩŶ-Ȕ ȖüϹḰɏǳÑǚǣƅά໐ť1ՈňϬ4ƅۘऺǕɴඓΝ Ēˊ⒲LôκˎiƏƋŦʦĝ¡ƂȐ˾ 2009 ˝Ethical ՈⒸ̱ᯬ¾ඊྡྷ Ởẋራ̵ᖏՁ issues in animal experimentation˵ʩŦʦȎŨ͸/ǙĝǰŸ۔ᄳՈϹḰɏͩ Ḱɏ̓ ˍᢆȓḰɏඊྡྷՈ⓷ɳᜬẂɋʺ¤[26] ╓؄ȕͧrⒸՈł⑃ ƚƫƫ᪱NσʗϏƉȼˍFĝhȗ^NσʗϏƉȼ ໐ɌɴϦʺʌᜬẂᱏÃ ᜬhTERTǳÑᝯň&ņএࣷǎᜬ FĆuȼˍFǓ͇ĝˍF9$ǒ(į·ʴǏġjļf ~ẂֲՈȖüՈ1࣏ඊྡྷ[27]4ǪƅǒɀۘऺǕɴ ϬϹृƈͩ ɡɪˋΡ~ȆŮɵi͸ΛˍFǡõƍƟĝļ]˝įˋȻΡ űƍƟɪƕΝDKƯϟ$ĜƤ0˲ĝƲ ḰɏȖü ǕɴֲՈȖüٰrᜬẂɋʌz࿆ᯬĿḰɏ5ͩ ŀ ΓȔɡǡõˍF üǸϹḰɏͩǳň&P੄řǳᜐ3ƅάՈ5ͩ[28-31]4Ĳ« ̚ˌ ΞŁẟPǹȴʌḰɏƣÏඊྡྷՈḰɏɋ ֲQ«PȨǣ ň້̌ɻģĳȔŀƋΡ~˼ǈ̧ĝȒɻƏ˫ĝ^Ƃ ͑ȆᩬՈ⒲L4 ̓ɡmüóQȒɻƏÇΛĝ°ûȕ͛ļ(Gģαƀ·ļÿ) 3.3 ôκΡ~Bˌĝ̵f ǒɀඓhTERTḰɏ๎ၠⒸ βǳΛϊcĶuʩʷşLáǫ̂˼̏ͦKæ4ě˺( 4ɬȒ ¦๎ၠⒸ̱ᯬ۔ᯬ¾ඊྡྷȒࢿΉ͑சɳ҉µƧ̱̓ ¾ඊྡྷࢿΉඈʐhTERTæ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈᜄச 4 ǆ໇ǲ References ˈ¿^சɳ҉ඈɨṇϦɴº[┑ ໐hTERTæ¦๎ၠⒸ [1] 5Ƶ,ŬᲳ,ƿࢄ·,਍.¦๎ၠⒸ̱ᯬ¾ඊྡྷՈ ࡿ3ȕͧǎℸǎ [J].Ἱ5Ŀƚ┦ƚĉ,2009,32(2):234-236. ̱ᯬ¾ඊྡྷࢿΉඈՈाွᜄசˈ¿ǌĺz¦๎ၠⒸ̱ᯬ¾ [2] Hou Y, Huang Q, Liu T, et al. Human amnion epithelial cells ඊྡྷࢿΉඈ ό:ȉẕȭ+ඈˈ¿4¦๎ၠⒸ̱ᯬ¾ඊྡྷ can be induced to differentiate into functional insulin- ࢿΉඈʐhTERTæ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉඈՈᜄϚྴˏ producing cells. Acta Biochim Biophys Sin. 2008;40(9):830- ௤ˈ¿^சɳ҉ඈɨṇ ţƅʺ¤ óྴှ҉ǃ࣏òλḿ4 839. [3] Okere B, Alviano F, Costa R, et al. In vitro differentiation of ᜄசȋ۔ǳᢅඓhTERTϹḰɏͩࢿΉȒ ȭƯẟசɳ҉̓ human amniotic epithelial cells into insulin-producing 3D :3ྴˏ௤ˈ¿ՈΝǜ3ྴှ҉ǃ࣏òՈλḿţƅṇ̓z spheroids. Int J Immunopathol Pharmacol. 2015;28(3):390-402. 4RT-PCR̼ϟ඗ȘǕɴhTERTæ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿ [4] Ekser B. Clinical xenotransplantation: the next medical Ήඈń750 bpǳᢅĽŎɳǝr ໐ȭ+ඈ3¦๎ၠⒸ̱ᯬ revolution. Lancet. 2012;379(9816):672-683. [5] Bouffi C, Bony C, Courties G, et al. IL-6-dependent PGE-2 ¾ඊྡྷࢿΉඈ3சɳ҉ඈ̼ϟƦᢅĽŎɳǝrᜬ\ secretion by mesenchymal stem cells inhibits local inflammation hTERT mRNAՈᜬẂ ȴ߾r̲ļɳhTERTȖüՈḰɏǳ in experimental arthritis. PLoS One. 2010;5(12):14247. ÑƯẟ¦๎ၠⒸ̱ᯬ¾ඊྡྷՈhTERTᜬẂ4ǒɀỞẋᆓ [6] Kristien E, Reekmans, Praet J, et al. Clinical potential of intravenous neural stem Cell delivery for treatment of Ƥஂæö൦ɏᄶᢆȓ4hTERTæ¦๎ၠⒸ̱ᯬ¾ඊྡྷࢿΉ neuroinflammatory disease in mice? Cell Transplantation. ඈྴှ҉ǃ࣏òºλḿ ȴ߾ඓhTERTϹḰɏ๎ၠⒸ̱ 2011;20(6):851-869.

ISSN 2095-4344 CN 21-1581/R CODEN: ZLKHAH 5831

[7] Hu J, Yu X, Wang Z, et al. Long term effects of the [22] Izumi M, Pazin BJ, Minervini CF, et al. Quantitative implantation of Wharton's jelly-derived mesenchymal stem comparison of stem cell marker-positive cells in fetal and term cells from the umbilical cord for newly-onset type 1 diabetes human anmion. J Reprod Immunol. 2009;81(1):39-43. mellitus. Endocr J. 2013;60(3):347-357. [23] Robb WB, Condron C, Moriarty M, et al. Taurine attenuates [8] Dufrane D, Goebbels RM, Gianello P. Alginate radiation-induced lung fibrosis in C57/Bl6 fibrosis prone mice. macroencapsulation of pig islets allows correction of Ir J Med Sci. 2010;179(1):99-105. streptozotocin-induced diabetes in primates up to 6 months [24] Christofidou-Solomidou M, Tyagi S, Tan KS, et al. Dietary without immunosuppression. Transplantation. 2010;90(10): flaxseed administered post thoracic radiation treatment 1054-1062. improves survival and mitigates radiation induced [9] Oghiso Y, Yamada Y. 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